Acute And Sub Acute Toxicity Studies Of Gandhaga Chunnam On Swiss Albino Rats

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RESEARCH ARTICLE

ACUTE AND SUB ACUTE TOXICITY STUDIES OF GANDHAGA CHUNNAM ON SWISS ALBINO RATS

*1

_{Om Sakthi, T.,}

2

_{Mohamed Musthafa, M. and}

3

_{Mohamed Hussain, M.}

1

_{PG Scholar, PG Department of Sirappu Maruthuvam, Govt.Siddha Medical College, Chennai, Tamil Nadu, India}

2

_{H.O.D, PG Department of Sirappu Maruthuvam, Govt.Siddha Medical College, Chennai, Tamil Nadu, India}

3

_{UG Scholar, Plant Biology and Bio-Technology, Loyola college, Chennai, Tamil Nadu, India}

ARTICLE INFO ABSTRACT

Psoriasis is a chronic, non- infectious skin disease affects both sexes. Gandhaga Chunnam (GC) has been employed as a traditional remedy for psoriasis which is a siddha herbo-mineral formulation contains gandhagam (sulphur) in its calcinated form. GC is further evaluated for toxicological effects. Acute and sub acute toxicity studies with GC were carried out on Swiss albinorats. During acute toxicity study there were no any adverse effects found in the general behavior and no mortality at any dose level given(2000 mg/kg/b.wt). In sub acute toxicity study in the dose of(200,400 mg/kg/b.wt) didn’t cause any changes in haematological & bio –chemical parameters like leucocyte count, free fatty acid, plasma and urine, creatinine level. Further histo-pathological examination of vital organs should normal suggesting no morphological disturbances. It can be considered that GC is safe and non-toxic.

Copyright © 2014 Om Sakthi et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

INTRODUCTION

Psoriasis is an auto immune disease characterized by well defined, slightly raised, dry erythematous macules with white

silvery scales with typical extensor in distribution. (Behl 1st

Edition 1962, Reprint 2007, 2009, 2011.In Indian systems of

medicine most practitioners formulate and dispense there own recipes. GC herbo-mineral formulation mainly consists of gandhagam. Honey was used as an vehicle to promote there action. Gandhagam (sulphur) has been reported to have

Astringent action (Thiyagarajan et al., 1952). Sulphur can

maximize skin health and it act as essential dietary mineral for both skin health and overall wellness of the body. Deficiency of this mineral in our body leads to progression of inflammatory skin diseases (Pub med: The use of Sulphur for

Skincare July 23rd, 2010 / by Sarah Terry.

www.livestrong.com / articles / 18286) In siddha literature Gandhagam is indicated for skin disorders, veneral diseases, urinary disorders and general debility (Thiyagarajan 2006). Coconut oil is used as a skin moisturiser (Murugesa mudhaliyar 1936). Egg white has an emulsent action (Gunapaadam thathu and jeeva vaguppu). But the proper toxicity of GC has not been studied earlier. So this research article is useful to evaluate the acute and sub acute toxicity of herbo mineral formulation GC in laboratory animals.

*Corresponding author: Om Sakthi, T.

PG Scholar, PG Department of Sirappu Maruthuvam, Govt.Siddha Medical College, Chennai, Tamil Nadu, India.

MATERIALS AND METHODS

Gandhaga chunnam has been prepared by the following method as per the text book (Anooboga vaidhya navaneetham

3rd Edition, 1st Print 1995, 3rd Print 2006)

Ingredients

1. Gandhagam(Sulphur) – 1400gm (40 palam)

2. Coconut oil – 2800gm (80 palam)

3. Egg white – Quantity sufficient.

4. Cucumber – 4 in number

Purification of drugs

Take large pieces of nellikkai Gandhagam (Sulphur) and embedded it in 3 to 4 long cucumber. Wrap the cucumber with thread. Heat the coconut oil in a pot and put that wrapped cucumber until the threads become charred. Allow it to cool and stored in a container.

Preparation method

Then the purified Gandhagam (Sulphur) is grounded with egg white for 3 hour (1 saamam) and made into pills of thetrankottai seed size (500mg) and dried in sunshade. After that the pills are placed inside the egg shell and it is covered by

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International Journal of Current Research

Vol. 6, Issue, 10, pp.9298--9303, October,2014

INTERNATIONAL JOURNAL OF CURRENT RESEARCH

Article History:

Received 10th_{July, 2014}

Received in revised form

15th_{August, 2014}

Accepted 04th_{September, 2014}

Published online 25th_October,₂₀₁₄

Key words:

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3 layers of mud pasted cloth (seelaiman) and are subjected for incineration (pudam) with 15 palam earu (cow dung cake – 525gm). Then the content is cooled and triturated in a stone mortor and then stored.

Dose: 65mg twice a day.

Adjuvant: Honey.

Duration: 48 days.

Aim: To evaluate the acute and sub acute toxicity of the siddha

drug ‘Gandhaga chunnam’.

Acute Toxicity study: OECD- 423 guidelines (Schlede

et al.,1992; Schlede et al.,1994)

Acute toxicity study for GC was carried out as per OECD guidelines423. Healthy female rat weighing 200-250 gms were divide into three groups. The animals were housed under standard conditions and room temperature was controlled. All animals were fed with standard rat pelleted diet. Studies carried out at 3 female rat under fasting condition. Signs of toxicity

was observed for every one hour for 1st 24 hours and every day

for about 14 days to observe any behavioural changes, physiological activities and mortality. Observations include changes in skin colour, eyes, mucus membrane, respiratory, circulatory, autonomic nervous system were systematically recorded. Study has got approval from the Institutional animal ethics committee (IAEC)(Approval No: IAEC/XXXIX/08/ CLBMCP/2013). GC was administered orally at doses of (50,100, 250,500,1000,2000 mg/kg/b.wt). No signs of toxicity was observed.

Sub Acute Toxicity study: (OECD guidelines 407) (Schlede

et al., 1992)

In sub acute toxicity study swiss albino rats of either sex weighing 220-250 gms were divided into three groups of 6 animals each (three male and three female) and were housed under the same conditions as described above. GC were administered for 28 days at doses of 200 and 400 mg/kg/b.wt respectively. The animals were observed daily for behavioural changes and other signs of sub acute toxicity. Toxic manifestations and mortality were monitored daily and body weight changes are recorded every 7 days till the end of study. The weight of each rat was recorded on day 0 and weekly thought the course of the study. Food and water consumption

per rat was calculated. On 29th day, animals were fasted for 18

hours, slightly anesthetized with ether and blood samples were collected from retro-orbital plexes.

Clinical parameters checked

Group Day

Body Weight Normal

Assessments of posture Normal

Signs of convulsion Limb paralysis Normal

Body tone Normal

Lacrimation Absence

Salivation Absence

Change in skin color No significant color change

Piloerection Not observed

Defecation Normal

Sensitivity response Normal

Locomotion Normal

Muscle grip ness Normal

Rearing Mild

Urination Normal

Effect of GC on Haematological analysis

The blood collected in one test tube and then added with EDTA and add heparin for immediate analysis of Haematological parameters (RBC count, total WBC count, differential white blood cell count, platelet count and haemoglobin) by semi auto analyser method. Blood collected in another tube without heparin was centrifuged at 4000 rpm

at40 c for 10 minutes to obtain serum and then stored at 200 c

for analysing of bio chemical parameters like creatinine, triglycerides, cholesterol and glucose by using standard methods.

RESULTS

Acute Toxicity study

There were no mortality and no behavioral changes, toxicity observed after oral administration of GC upto the dose level 2000 mg/kg/b.wt in rats.

Sub acute Toxicity study

There was no significant difference between the control and GC treated groups. No lethality was recorded for any dose upto

the maximum of 400 mg/kg during the 28day period of testing.

Statistical analysis

Effect of GC in animal model was effective and highly significant.

Grouping Total RBC

Count(x106 µ l)

Total WBC

Count(x103 µ l)

Platelet

count(x103 µ l)

PCV (%) MCV (fl) MCH (pg) MCHC (g/dl) Blood Sugar ® (mg/dl)

BUN (mg/dl) Control

Mean 7.667 7.333 567.7 45.5 56.17 21.67 35 76.5 16.5

SD 1.506 0.8165 5.61 3.271 2.317 4.227 4.49 3.146 2.811

SE 0.6146 0.3333 2..29 1.335 0.9458 1.726 1.751 1.285 1.1147

Low Dose

Mean 8.417 9.35 570.5 47.83 58.67 24.17 41.5 74.5 13.58

SD 1.009 1.193 2.074 2.563 4.082 4.708 1.975 3.834 1.855

SE 0.4118 0.487 0.8466 1.046 1.667 1.922 0.8062 1.565 0.7574

High Dose

Mean 8.583 6.883 564.2 42.67 55.5 20 45.67 71.33 14

SD 0.7859 1.234 4.916 3.777 3.619 2.366 3.882 3.615 3.347

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Effect of GC on lipid profile

Grouping Serum creatinine

(mg/dl)

Serum Total Colorestrol (mg/dl)

SerumTGL Level (mg/dl)

SerumHDL Level (mg/dl)

Serum LDL Level (mg/dl)

Serum VLDl (mg/dl) Control

Mean 0.8167 97.67 46.33 26.7 45.5 35.5

SD 0.07528 3.077 3.327 2.733 3.507 3.728

SE 0.03073 1.256 1.358 1.116 1.432 1.522

Low Dose

Mean 0.8167 107.2 49.67 26.83 46 34.5

SD 0.2714 2.563 4.131 3.251 4.336 3.45

SE 0.1108 1.046 1.687 1.327 1.77 1.408

High Dose

Mean 0.5333 102.5 41.83 32.5 42.33 35

SD 0.1633 4.183 2.848 2.51 3.502 4.05

SE 0.06667 1.708 1.167 1.025 1.43 1.653

Effect of GC on Body weight and Food consumption

Grouping Food (g/day/rat) Body weight (g)

CONTROL

MEAN 20.17 231

SD 1.329 9.839

SE 0.5426 4.017

LOW DOSE Food (g/day/rat) Body weight (g)

MEAN 28.5 231.3

SD 3.728 3.615

SE 1.522 1.476

HIGH DOSE Food (g/day/rat) Body weight (g)

MEAN 21.67 225.8

SD 2.944 4.834

SE 1.202 1.973

Effect of GC on Mortality rate of the study animals

Treatment Mortality observed for the duration of 1- 28 days

GROUP I – CONTROL NIL

GROUP II- LOW DOSE NIL

GROUP III- HIGH DOSE NIL

Effect of GC on organ morphology

Grouping Kidney Liver Heart lungs Spleen pancreas Brain Ovaries Testes

GROUP I CONTROL

Normal Normal Normal Normal Normal Normal Normal Normal Normal

GROUP II LOW DOSE

GROUP III HIGH DOSE

Histo pathological analysis of Sub-Acute toxicity study

Group Control Low Dose High Dose

Kidney Magnification: Low Power 10X

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High power 45X

Group Control Low Dose High Dose

High Power 45X

Heart

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Software: spss17 version

Test: Paired t test

Confidence Interval: 95%

Correlation coefficient (r): 0.736

Mean difference ± SD: 17.07 ± 8.41

P Value (2 tailed): p<0.01.

Inference

Since the P value is highly significant (<0.01). Hence it is

concluded that the GC was effectiveand significant.

DISCUSSION

In Acute toxicity study there was no any mortality observed upto the maximum level of 2000 mg/kg/b.wt of GC administered orally, which the single high dose recommended

by OECD guidelines – 4235 for testing acute toxicity. In that

GC doesn’t cause any acute toxicity. The changes in body weight have been used as an indicator of adverse effect of GC and chemicals (Ecobichon 1997). Since there were no changes in animal behaviour, body 7 organ weights at all doses of treated rats when compared to the control group. The present results reveal that at the oral doses administered GC is

PATHOLOGIST REPORT

Sample Observation

Kidney Pattern of nephrons and arteries appears normal , no Sevier

abnormalities in all three groups

Heart Nucleus appears dense and intense and myocardial fiber length appears

normal.

Liver No signs of necrosis or cirrhosis were observed in all the groups.

Lumen of hepatic veins appears normal.

Brain Neuronal integrity appears normal in all the groups and also

no considerable observation of signs of edema or degeneration

High power 45X

Brain

Magnification: Low power 10X

High Power 45X

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toxic. Histo pathological studies of skeletal organs (Heart, Liver, Kidney, and Brain) from both treated and control animals shows normal architecture, suggesting no detrimental changes and morphological disturbances caused due to the administration of GC for 28 days.

Conclusion

GC can be considered safe and didn’t cause either any lethality or adverse changes with general behavior of rats in aute toxicity study upto the dose of 2000 mg/kg b.wt and also there was no observed detrimental effects caused by GC (upto 400 mg/kg b.wt) in sub acute study rat model. Further the above

results substantiate the beneficial and enhanced

pharmacological effects of the herbo mineral formulation GC.

Acknowledgement

I am very thankful for Dr.M .Mohamed musthafa, M.D (S)., HOD, P.G Sirappu maruthuvam department, GSMC Chennai, Tamilnadu, India, for his guidance in this study and publishing it.

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